Process and device for preparation of cell samples for cytological tests

ABSTRACT

Process for preparing cell samples for cytological tests of exfoliated cells, in which the cell sample is in the form of a slurry in a physiological solution, which is introduced into a first chamber (5) provided with a wall (2) with a large number of holes with a cross section of 10-100 μm and communicating with a second chamber (6). The solution is pressed under the influence of a pressure differential between the first (5) and the second (6) chamber through the wall (2) with holes. The material for the cell sample is taken from the solution in the second chamber (6) for smearing on a slide. A suitable device for carrying out the process is also described.

The present invention relates to a process for preparation of cellsamples for cytological tests of exfoliated cells, and a device forcarrying out the process.

Cytological tests of cell samples are now quite prevalent and haveproved to be an exceptional means for diagnosing cell changes in thearea about the portio-cervix and endocervical canal. Such tests areperformed on large numbers of women and not infrequently on entire agegroups of women within a district for example.

There are also tests of cell samples taken from other portions of thebody done regularly at the cytological laboratories, but not other testis as frequently done as the cervical smear. A cell sample taken fromthe cervical canal reveals cell changes with a reliability of about 95%,and of these 20-25% are precancerous. A positive response to the cellchange test usually results in an operation, a so-called scraping or useof a cone instrument. The results of such treatment are good and wellwarrant extensive testing of healthy women in certain age groups.

The samples are taken with a curette or similar instrument by scrapingcells from the mouth of the cervix. It is of course very important thatany cell changes be represented in the sample. There are a number ofdifferent types of sampling devices which provide acceptable samples.The samples are taken by inserting a speculum into the vagina whereafterthe sampling instrument is inserted and the scraping is done.

The sample material must then be protected from destruction while beingtransported to the cytological laboratory, suitably by immersion in afixing solution.

After the sample has arrived at the laboratory it is prepared by dyeingand is inspected in a microscope. The presence of atypical cells isnoted and reported. The examination under the microscope and theevaluation requires exceptional attentiveness and is time-consuming, andis considered to be quite demanding work. In 1980 each test cost between50 and 100 Swed.Kronor.

Consequently, intensive development work is in progress to simplify andmake less expensive sampling, sample preparation and sample evaluation.In order to simplify the evaluation of the samples under the microscope,a number of systems have been developed for automatic evaluation ofcells as normal or atypical. Algorithms have been formulated forautomatic evaluation and the development of commercial systems is fairlyfar along.

It is thus possible with reasonably good accuracy to determine ifatypical cells are present in a cell sample, if the cell sample has beenprepared so that a substantial portion of the cells are free cells onthe slide. Under certain conditions, a human evaluator can alsorecognize atypical cells in clumps of cells, however.

This is not the case with automatic examination of a cell sample andthus it is of crucial importance in this case to have as many free cellsas possible. Samples have been prepared previously with a conventionalsyringe, for example, provided with a cannula with a diameter of 500 μm,by alternatingly sucking up and expelling the slurry of scraped cells.This produced cell samples which could be read by an experienced humanexaminer. In order to be read automatically, the sample preparation mustbe improved so that the majority of the cells in the sample are freecells.

The purpose of the present invention is thus to provide an effectiveprocess for sample preparation of cell samples and a device for carryingout the process.

The new process is essential for establishing a functioning system forautomatic cell testing and facilitating substantially the non-automatedevaluation of cell samples.

The new process, intended for the preparation of cell samples forcytological testing of exfoliated cells, in which the cell sample is inthe form of a slurry in a fixing solution, is characterized in that theslurry is introduced into a first chamber provided with a wall with alarge number of holes with a cross section of 10-100 μm and incommunication with a second chamber and that said solution, under theinfluence of a pressure differential, is made to pass through the wallprovided with holes one or more times, and that the material from thesecond chamber is used for the preparation of smears on a slide.

It is preferable that the cross section of the holes being 10-40 μm. Ifthe cross section is less than 5 μm, the results will not bereproducible, and if the cross section is more than 100 μm, the resultsof the test preparation will be less than satisfactory.

After one or more passages through the wall provided with holes, thesolution is allowed to settle in the second chamber and a smear on aslide is made from the sedimented material.

It has been shown to be suitable to use a pressure differential betweenthe first and the second chambers of 50-1000 kPa to press the slurrythrough the wall provided with holes.

The pressure differential is suitably established by pressing a plungerinto a cylindrical cavity, said cavity communicating with the firstchamber.

The slurry suitably contains a cell dissociating agent, which can be forexample hyaluronidase, chymotrypsin or 1,4-dimercapto-2,3-butane diol.Fixing agent, such as 50-95% ethanol is also included and suitably alsoa small amount of sodium hydroxide.

A device for carrying out the new process consists of a first and asecond chamber separated by a wall, provided with a large number ofholes with a cross section of 10-100 μm. It is also provided with meansfor establishing a pressure differential between the first and thesecond chamber. Furthermore, it is arranged to make it possible tointroduce slurry into the first chamber and to extract finished,possibly sedimented sample material from the second chamber. The devicecan consist of a first tube, closed at one end and into which a secondtube has been partially inserted, which at its inserted end is providedwith a large number of holes and is essentially sealingly connected tothe first tube. A movable plunger is arranged in the second tube and canbe moved reciprocally there. Said plunger can also be inserted andremoved from the second tube for introducing the slurry into the secondtube.

A preferred embodiment of the invention will be described in more detailwith reference to the accompanying drawing.

The device consists of three parts: a tube 1 provided with a wall 2 witha plurality of holes, a plunger 3 and an outer closed tube 4. The tube 1with the wall 2 and the plunger 3 defines the first chamber 5. The tube1 can be inserted into the outer tube 4 suitably so as to seal againsteach other. The second chamber 6 is defined in the tube 4 by the lowerend of the closed tube and the wall tube provided with holes. The outertube 4 is suitably a centrifuge tube with a pointed lower end 7. Thetube 1 and the plunger 3 are suitably made as a hypodermic syringe witha flange 8 and a pressing surface 9 as well as wings 10 for centeringthe movement of the plunger in the tube 1 when pressure is exerted onthe pressure surface 9. The plunger 3 is suitably provided with a rubberpacking 11 to provide a better seal between the plunger 3 and the tube1.

The device shown is intended for manual sample preparation, the plunger3 first be removed from the tube 1 and the sample slurried in thephysiological fluid being poured down into the tube 1. The plunger 3 isinserted and then pressed down through the tube 1. The solution willthen be pressed through the wall with holes, suitably represented by awire-mesh or a net of artificial fibers with a cross section diameter of10-100 μm. The tubes 1 and 4 can then be turned over and the plunger 3drawn out, thereby sucking the solution through the wall with holes,whereafter the liquid is again pressed with the plunger through the wallwith holes. The process is suitably repeated a number of times,whereafter the liquid is sedimented or centrifuged in the tube 4. Thematerial for the smear can then be taken from the sedimental materialthus providing a smear which has free cells to a large degree and whichis suitable for automatic evaluation.

The person skilled in the art should have no difficulty conceivingautomatic units in which the sample preparation can be done completelyautomatically even if we have not described here an example of such adevice.

I claim:
 1. Process for preparation of cell samples for cytologicaltests of exfoliated cells, in which the cell sample is in the form of aslurry in solution, characterized in that the slurry is introduced intoa first chamber provided with a wall with a large number of holes with across section of 10-100 μm and in communication with a second chamber,and that said solution under the influence of a pressure differentialbetween the first and the second chambers is made to pass through thewall provided with holes one or more times, and that the material fromthe second chamber is used for the preparation of smears on a slide. 2.Process according to claim 1, characterized in that the cross section ofthe holes is 10-40 μm.
 3. Process according to claim 1, characterized inthat a pressure differential of 50-1000 kPa is used to press the slurrythrough the wall provided with holes.
 4. Process according to claim 1,characterized in that the liquid is allowed to sediment in the secondchamber and that smears are made from the sediment material.
 5. Processaccording to claim 1, characterized in that the pressure differential isestablished by pressing a plunger into a cylindrical cavity, said cavitybeing in communication with the first chamber.
 6. Process according toclaim 1, characterized in that the slurry contains a cell dissociatingagent and a fixing agent.
 7. Process according to claim 6, characterizedin that the cell dissociating agent consists of hyaluronidase,chymotrypsin or 1,4-mercapto-2,3-butane diol and the fixing agent of50-95% ethanol and that the slurry contains a small amount of sodiumhydroxide.
 8. Process according to claim 1, characterized in that theslurry is made to pass through the wall by alternating pressure andsuction effect.